大动物胸腺修饰诱导同种心脏移植免疫耐受的实验研究

Induction of Allogenic Cardiac Transplantation Tolerance by Intrathymic Inoculation of Donor Spleen Cells of Pigs

使用胸腺修饰和γ射线全身照射方法预处理受体猪 ,来评估该诱导免疫耐受方法用于大动物同种异体心脏移植的可能性。动物分组 :空白组不加处理行异位心脏移植 ;对照组受体单用 2Gyγ 射线全身照射 ;实验组受体全身照射加胸腺内注射供体脾细胞 ,两组预处理后 ,观察外周血中淋巴细胞及其亚群CD4、CD8水平的变化以及使用混合淋巴细胞反应(MLR )来了解其免疫功能的相关改变。预处理后 14d ,行异位心脏移植 ,观察移植物生存时间 ,并检测术后IL 2、IL 10水平变化。接受照射 4 8h后淋巴细胞较照射前显著减少 (P <0 0 5 ) ,流式细胞仪计数CD4、CD8水平也明显下降 ,但CD4下降更为明显 ;实验组受体胸腺修饰后第 2周 ,混合淋巴细胞反应刺激效应有所下降 (P <0 0 5 ) ,而单纯照射组预处理前后无显著差异 (P >0 0 5 )。实验组供心存活期 (MST 7 4d )较空白组 (MST 4 2d )和照射组 (MST 4 6d )明显延长 (P <0 0 5 ) ,而空白组与单纯照射组比较无显著差异 (P >0 0 5 )。实验组移植后 1～ 5dIL 2的水平明显低于对照组和空白组 (P <0 0 5 ) ,IL 10水平三组无明显差异 (P >0 0 5 )。实验组预处理后 ,机体产生了一定程度的免疫抑制 ,但并未获得在小动物实验中所获得的所谓“永久性”免疫耐受 。

To search for the possibility to induce allogenic cardiac transplantation tolerance in pigs by intrathymic inoculation of donor spleen cells and the pretreatment of the recipient pigs with γ ray whole body irradiation,animals were divided into 3 groups,in which the blank group (A) was without any treatment except the allogenic cardiac transplantation,control group (B) was pretreated with 2 Gy γ ray irradiation and experimental group (C) was pretreated with whole body irradiation and inoculated intrathymically with donor spleen cells. In these two pretreament groups,the changes of the peripheral blood lymphocytes,the levels of CD4,CD8 T cell subsets and mixed lymphocyte reaction (MLR) were observed to test any alterations of immune functions. The allogenic cardiac transplantation was performed 14 days after pretreatment,and transplant survival times observed with simultaneous testing of the levels of IL-2 and IL-10. It was found that after 48 hours of irradiation,the number of lymphocytes was significantly decreased in comparison with that before irradiation,and the levels of CD4 and CD8 cells also reduced as determined by flow cytometry analysis,the reduction in the CD4 cells being even more prominent. In the experimental group 2 weeks after intrathymic inoculation of spleen cells in recipient pigs,the stimulatory effect in MLR showed a tendency to reduce,while that with single irradiation showed no difference before and after pretreatment. The survival time of the donor cardiac grafts in the experimental group (MST 7.4 days) was clearly prolonged in comparison with those of blank group (MST 4.2 days) and control group (MST 4.6 days),but there was no significant difference between blank and single irradiation group. The level of IL-2 after 1～5 days of grafting was significantly lower than those of the control and blank groups,but that of IL-10 among these three groups showed no difference. These experimental results indicate that the intrathymic inoculation with donor spleen cells and pretreatment with whole body irradiation of the recipient out bred pigs could induce a transient immune suppression,but not attain to the so called permanent immune tolerance as observed in small animals. There is a close relationship between the level of IL-2 and the development of rejection reactions.